HIV-1 latency, which allows the virus to persist in the presence of antiretroviral therapy, is likely the major hurdle to overcome towards the development of a curative therapy for HIV-1 infection. While it is clear that the latent HIV-1 reservoir needs to be eradicated, it is unclear how this can be achieved. At the molecular biology level, current HIV-1 research focuses on the idea that molecular control of latent HIV-1 infection must be different from the control of inducible cellular genes and is characterized by the presence of a restrictive chromatin environment at the viral LTR. But how such a restrictive chromatin environment can form when the latent virus is integrated into actively expressed host-genes is unclear. How can paused RNAP II be found at the latent viral LTR, when a complex restrictive chromatin structure supposedly shields the LTR? This focus on a restrictive chromatin environment as the molecular control mechanism for latent HIV-1 infection translates into drug discovery and clinical application, where HDAC inhibitors are viewed as the holy grail of HIV-1 reactivation strategies. However, upon closer review of the literature, HDAC inhibitors, which are clinically pursued as HIV-1 reactivating agents, do not show convincing efficacy, neither in many models of latent infection, nor in ex vivo experiments or clinical studies. The possible exception is SAHA, clinically approved as the HDAC inhibitor vorinostat. SAHA exhibits some HIV-1 reactivating capacity in most experimental systems, including ours, but SAHA was initially developed as a highly potent cell-differentiating agent, using HMBA, another cell differentiating agent that reactivates latent HIV-1 as a structural template. To this end, we here report that a previous drug screen for HIV-1 reactivating drug combinations revealed that a panel of FDA-approved drugs or compounds with reported cell-differentiating capacity including dactinomycin, aclacinomycin, cytarabine and aphidicolin prime latent infection for reactivation by low-level activation. We hypothesize that cell-differentiating drugs can act to prime latent HIV-1 infection for reactivation by synergistic activators, which by themselves only exert a minimal activating effect. Thus the two objectives of the application are (i) to identify synergistic activators that trigger system-wide reactivation in combination with the priming drugs and (ii) to describe how differentiating drugs alter the cellular transcription factor profiles and signal transduction pathways to prime latent HIV-1 infection for reactivation. The goal of the application is to identiy multi-drug combinations that target latent HIV-1 infection at several levels of molecular control t trigger reactivation. This will be achieved through drug screening or by rational selection of compounds/drugs that will be enabled as we gain increasingly detailed insights into how the identified drugs alter the cellular control over the latent HIV-1 infection events.